REAGENT or RESOURCE	SOURCE	IDENTIFIER
Antibodies
Anti-FLAG M2 Magnetic Agarose Beads	Sigma-Aldrich	Cat# M8823; RRID: AB_2637089
GFP-Trap Agarose Beads	ChromoTek	Cat# gta-20; RRID: AB_2631357
Bacterial and virus strains
Escherichia coli strain : OP50	Caenorhabditis elegans Center	OP50
Chemicals, peptides, and recombinant proteins
Cas9-NLS protein TrueCut V2 (5mg/mL)	Invitrogen	Cat#A36498
TRI Reagent	Invitrogen	Cat# AM9738
Turbo Dnase	Invitrogen	Cat#AM2238
HALT Protease Inhibitors	Thermo Fisher Scientific	Cat#78425
RiboLock RNAse Inhibitor	Thermo Fisher Scientific	Cat#EO0381
75 mM Bio-11-UTP	Invitrogen	Cat#10269614
T4 Polynucleotide Kinase	New England Biolabs	Cat#M0201L
DreamTaq DNA polymerase	Thermo Fisher Scientific	Cat#11883813
pronase E	Sigma	Cat#7433-2
Critical commercial assays
Qubit Fluorometer High Sensitivity dsDNA assay kit	Thermo Fisher Scientific	Cat#Q32851
NextSeq 500/550 High Output v2 kit 75 cycles	Illumina	Cat# FC-404-2005
NEBNext Ultra II Directional RNA Library Prep Kit for Illumina	New England Biolabs	Cat#E7760S
SuperScript IV Reverse Transcriptase	Thermo Fisher Scientific	Cat#18090010
NEBNext Ultra II Q5 Master Mix	New England Biolabs	Cat#M0544L
M-MLV reverse transcriptase	Invitrogen	Cat#28025013
Deposited data
All sequencing data	This study	GSE157319
Original polyUG PCR gel images	This study	DOI:10.17632/tnmxzg8wjj.1
Experimental models: Organisms/strains
C. elegans strain: wild isolate (Bristol)	CGC	N2
C. elegans strain: piwi(n4357) I	CGC	SX922
C. elegans strain: hrde-1(tm1200) II	CGC	FX1200
C. elegans strain: rde-3(ne3370)	CGC	WM286
C. elegans strain: ggSi1[hrde-1p::3xflag::gfp::hrde-1] II	S. Kennedy lab	YY584
C. elegans strain: piwi(gcp034[STOP]) I; ggSi1[hrde-1p::3xflag::gfp::hrde-1] II	This study	MHE109
C. elegans strain: piwi(gcp035[STOP]) I; ggSi1[hrde-1p::3xflag::gfp::hrde-1] II	This study	MHE110
C. elegans strain: drh-3(ne4253) I; ggSi1[hrde-1p::3xflag::gfp::hrde-1] II	This study	MHE138
C. elegans strain: mut-16(pk710) I; ggSi1[hrde-1p::3xflag::gfp::hrde-1] II	This study	MHE141
C. elegans strain: rde-3(ne3370); ggSi1[hrde-1p::3xflag::gfp::hrde-1] II	This study	MHE194
C. elegans strain: gfp::prg-1; flag::mcherry::glh-1 I	HC. Lee lab	HCL125
C. elegans strain: csr-1(gcp027[mCherry::3xflag::1ha::csr-1]) IV; glh-1(sam24[glh-1::gfp::3xFlag]) I	This study	MHE79
C. elegans strain: piwi(gcp046[mCherry::prg-1]) I; mut-16(cmp3[mut-16::gfp::3xFLAG + loxP]) I	This study	MHE147
C. elegans strain: mut-16(cmp3[mut-16::gfp::3xFLAG + loxP]) I; csr-1(gcp027[mCherry::3xflag::1ha::csr-1] IV	This study	MHE 93
C. elegans strain: piwi(gcp038) I; mut-16(cmp3[mut-16::gfp::3xFLAG + loxP]) I; csr-1(gcp027[mCherry::3xflag::1ha::csr-1] IV	This study	MHE117
C. elegans strain: csr-1(gcp017[csr-1::3xFLAG::1HA])IV	G. Cecere lab	MHE27
C. elegans strain: csr-1(tm892)/nT1 [unc-?(n754) let-? qIs51] (IV;V)	G. Cecere lab	MHE 20
C. elegans strain: csr-1(gcp010)[D769A]/nT1 [unc-?(n754) let-? qIs51] (IV;V)	G. Cecere lab	MHE 21
C. elegans strain: glh-1(sam24[glh-1::gfp::3xFlag]) I	D. Updike lab	DUP64
C. elegans strain: glh-1(sam24[glh-1::gfp::3xFlag]) I; csr-1(tm892)/nT1 [unc-?(n754) let-? qIs51] (IV;V)	This study	MHE 48
C. elegans strain: glh-1(sam24[glh-1::gfp::3xFlag]) I; csr-1(gcp017[csr-1::3xFLAG::1HA];gcp022[D769A]/nT1[unc-?(n754) let-? qIs51] (IV;V)	This study	MHE 49
C. elegans strain: csr-1(gcp043[αN::3xFLAG::HA::CSR-1]) IV	This study	MHE131
C. elegans strain: ZK795.2(gcp064[zk795.2::3UTR_5boxb]) IV	This study	MHE186
C. elegans strain: ZK795.2(gcp066[zk795.2::3UTR_5boxb]) IV; csr-1(gcp043[αN::3xFLAG::HA::CSR-1]) IV	This study	MHE188
C. elegans strain: ZK795.2(gcp067[intronic_zk795.2::5boxb]) IV	This study	MHE191
C. elegans strain: ZK795.2(gcp067[intronic_zk795.2::5boxb]) IV; csr-1(gcp043[αN::3xFLAG::HA::CSR-1]) IV	This study	MHE197
C. elegans strain: hsp-90(gcp068[Phsp-90::αN::3xFLAG::mCherry] V	This study	MHE201
C. elegans strain: ZK795.2(gcp064[zk795.2::3UTR_5boxb]) IV; hsp-90(gcpp069[Phsp-90::αN::3xFLAG::mCherry]) V	This study	MHE202
Recombinant DNA
pJJR82 (EGFPˆSECˆ3xFlag)	Mike Boxem lab	Addgene plasmid # 75027; http://n2t.net/addgene:75027; RRID:Addgene_75027
pJJR83 (mCherryˆSECˆ3xFlag)	Mike Boxem lab	Addgene plasmid # 75028; http://n2t.net/addgene:75028; RRID:Addgene_75028
pFA6a-5BoxB-hphMX6 (5boxb)	Marc Bühler lab	(Bühler et al., 2006)
pRF4::rol-6(su1006)	(Mello et al., 1991)	N/A
Software and algorithms
Integrative Genomics Viewer	http://www.broadinstitute.org/igv/	RRID:SCR_011793
Custom scripts and workflows for sequencing analysis	https://gitlab.pasteur.fr/bli/bioinfo_utils	DOI:10.5281/zenodo.5720645
Deeptools	https://deeptools.readthedocs.io/en/develop/	RRID:SCR_016366
Fiji	https://fiji.sc/	RRID:SCR_002285
StackReg (Fiji plugin)	(Thévenaz et al., 1998)	http://bigwww.epfl.ch/thevenaz/stackreg/
Oligo melting Python package	https://zenodo.org/record/4593033	DOI:10.5281/zenodo.4593033
iFISH probe design (ifpd)	https://ggirelli.github.io/iFISH-probe-design/	DOI:10.5281/zenodo.5584634
ImJoy	https://imjoy.io/#/	RRID:SCR_020935
Analysis workflows for smFISH image quantifications	https://github.com/fish-quant/fq-imjoy	DOI:10.5281/zenodo.5718531
Source code to analyze RNA localization in C. elegans germlines	https://github.com/muellerflorian/cornes-rna-loc	DOI:10.5281/zenodo.5718481